Functional acylglycerides

ABSTRACT

The present invention relates to the field of human and animal nutrition, and in particular to certain novel compositions of conjugated linoleic acids (CLA). In particular, the present invention relates certain isomers of conjugated linoleic acids at either the SN1 and SN3 or SN2 positions of the acylglyceride molecule and another fatty acyl residue at the other of the SN1 and SN3 or SN2 positions of the acylglyceride molecule. These novel acylglyceride compositions containing conjugated linoleic acyl residues, medium chain fatty acyl residues, long chain fatty acyl residues, and ω3, ω6, and ω9 fatty acyl residues are efficacious as pharmaceutical compositions, animal feed additives, and human dietary supplements.

This application is a continuation of U.S. application Ser. No.09/989,835, filed Nov. 20, 2001 now U.S. Pat. No. 6,677,470, allowed.

FIELD OF THE INVENTION

The present invention relates to the field of human and animalnutrition, and in particular to certain novel compositions of conjugatedlinoleic acids (CLA). In particular, the present invention relates tocertain isomers of conjugated linoleic acids at either the SN1 and SN3or SN2 positions of the acylglyceride molecule and another fatty acylresidue at the other of the SN1 and SN3 or SN2 positions of theacylglyceride molecule.

BACKGROUND OF THE INVENTION

In 1978, researchers at the University of Wisconsin discovered theidentity of a substance contained in cooked beef that appeared toinhibit mutagenesis. The substance was found to be a mixture ofpositional isomers of linoleic acid (C18:2) having conjugated doublebonds. The c9,t11 and t10,c12 isomers are present in greatest abundance,but it is uncertain which isomers are responsible for the biologicalactivity observed. It has been noted from labeled uptake studies thatthe 9,11 isomer appears to be somewhat preferentially taken up andincorporated into the phospholipid fraction of animal tissues, and to alesser extent the 10,12 isomer. (Ha, et al., Cancer Res., 50: 1097[1990]).

The biological activity associated with conjugated linoleic acids(termed CLA) is diverse and complex. At present, very little is knownabout the mechanisms of action, although several preclinical andclinical studies in progress are likely to shed new light on thephysiological and biochemical modes of action. The anticarcinogenicproperties of CLA have been well documented. Administration of CLAinhibits rat mammary tumorigenesis, as demonstrated by Birt, et al.,Cancer Res., 52: 2035s [1992]. Ha, et al., Cancer Res., 50: 1097 [1990]reported similar results in a mouse forestomach neoplasia model. CLA hasalso been identified as a strong cytotoxic agent against target humanmelanoma, colorectal and breast cancer cells in vitro. A recent majorreview article confirms the conclusions drawn from individual studies(Ip, Am. J. Clin. Nutr., 66 (6 Supp): 1523s [1997]).

Although the mechanisms of CLA action are still obscure, there isevidence that some component(s) of the immune system may be involved, atleast in vivo. U.S. Pat. No. 5,585,400 (Cook, et al., incorporatedherein by reference), discloses a method for attenuating allergicreactions in animals mediated by type I or TgE hypersensitivity byadministering a diet containing CLA. CLA in concentrations of about 0.1to 1.0 percent was also shown to be an effective adjuvant in preservingwhite blood cells. U.S. Pat. No. 5,674,901 (Cook, et al.), incorporatedherein by reference, disclosed that oral or parenteral administration ofCLA in either free acid or salt form resulted in elevation in CD-4 andCD-8 lymphocyte subpopulations associated with cell-mediated immunity.Adverse effects arising from pretreatment with exogenous tumor necrosisfactor could be alleviated indirectly by elevation or maintenance oflevels of CD-4 and CD-8 cells in animals to which CLA was administered.Finally, U.S. Pat. No. 5,430,066, incorporated herein by reference,describes the effect of CLA in preventing weight loss and anorexia byimmune stimulation.

Apart from potential therapeutic and pharmacologic applications of CLAas set forth above, there has been much excitement regarding the use ofCLA nutritively as a dietary supplement. CLA has been found to exert aprofound generalized effect on body composition, in particularredirecting the partitioning of fat and lean tissue mass. U.S. Pat. No.5,554,646 (Cook, et al.), incorporated herein by reference, discloses amethod utilizing CLA as a dietary supplement in which pigs, mice, andhumans were fed diets containing 0.5 percent CLA. In each species, asignificant drop in fat content was observed with a concomitant increasein protein mass. It is interesting that in these animals, increasing thefatty acid content of the diet by addition of CLA resulted in noincrease in body weight, but was associated with a redistribution of fatand lean within the body. Another dietary phenomenon of interest is theeffect of CLA supplementation on feed conversion. U.S. Pat. No.5,428,072 (Cook, et al., incorporated herein by reference), provideddata showing that incorporation of CLA into animal feed (birds andmammals) increased the efficiency of feed conversion leading to greaterweight gain in the CLA supplemented animals.

Another important source of interest in CLA, and one which underscoresits early commercial potential, is that it is naturally occurring infoods and feeds consumed by humans and animals alike. In particular, CLAis abundant in products from ruminants. For example, several studieshave been conducted in which CLA has been surveyed in various dairyproducts. Aneja, et al., J. Dairy Sci., 43: 231 [1990] observed thatprocessing of milk into yogurt resulted in a concentration of CLA.(Shanta, et al., Food Chem., 47: 257 [1993]) showed that a combinedincrease in processing temperature and addition of whey increased CLAconcentration during preparation of processed cheese. In a separatestudy, Shanta, et al., J. Food Sci., 60: 695 [1995] reported that whileprocessing and storage conditions did not appreciably reduce CLAconcentrations, they did not observe any increases. In fact, severalstudies have indicated that seasonal or interanimal variation canaccount for as much as three fold differences in CLA content of cowsmilk. (See e.g., Parodi, et al., J. Dairy Sci., 60: 1550 [1977]). Also,dietary factors have been implicated in CLA content variation, as notedby Chin, et al., J. Food Camp. Anal., 5: 185 [1992]. Because of thisvariation in CLA content in natural sources, ingestion of prescribedamounts of various foods will not guarantee that the individual oranimal will receive the optimum doses to ensure achieving the desirednutritive effect.

Linoleic acid is an important component of biolipids, and comprises asignificant proportion of triglycerides and phospholipids. Linoleic acidis known as an “essential” fatty acid, meaning that the animal mustobtain it from exogenous dietary sources since it cannot beautosynthesized. Incorporation of the CLA form of linoleic acid mayresult in a direct substitution of CLA into lipid positions whereunconjugated linoleic would have migrated. However, this has not beenproven, and some of the highly beneficial but unexplained effectsobserved may even result from a repositioning of CLA within the lipidarchitecture at sites where unconjugated linoleic acid would not haveotherwise migrated. It is now clear that one source of animal CLA,especially in dairy products, comes from the biochemical action ofcertain rumen bacteria on native linoleic acid, first isomerizing thelinoleic acid to CLA, and then secreting it into the rumen cavity.Kepler, et al., J. Nutrition, 56: 1191 [1966] isolated a rumenbacterium, Butyrivibrio fibrisolvens, which catalyzes formation of9,11-CLA as an intermediate in the biohydrogenation of linoleic acid.Chin, et al., J. Nutrition, 124: 694 [1994] further found that CLA foundin the tissues of rodent was associated with bacteria, sincecorresponding germ-free rats produced no CLA.

While the free fatty acid forms of conjugated linoleic acid describedabove are suitable for some uses, what is needed in the art are forms ofconjugated linoleic acid tailored for particular purposes.

SUMMARY OF THE INVENTION

The present invention relates to the field of human and animalnutrition, and in particular to certain novel compositions of conjugatedlinoleic acids (CLA). In particular, the present invention relates tocertain isomers of conjugated linoleic acids at either the SN1 and SN3or SN2 positions of the acylglyceride molecule and another fatty acylresidue at the other of the SN1 and SN3 or SN2 positions of theacylglyceride molecule.

Accordingly, in some embodiments, the present invention providesacylglycerides having the following structure:

wherein R1 and R3 are acyl residues selected from the group consistingof 10,12; 9,11; 8,10; and 11,13 octadecadienoate and R2 is selected fromthe group consisting of long chain and medium chain fatty acyl residues.The present invention is not limited to acylglycerides containing anyparticular medium chain fatty acyl residue. Indeed, a variety of mediumchain fatty acyl residues are contemplated including, but not limitedto, residues of the following acids: decanoic acid, undecanoic acid,10-undecanoic acid, lauric acid, cis-5-dodecanoic acid, tridecanoicacid, myristic acid, and myristoleic acid. The present invention is notlimited to acylglycerides containing any particular long chain fattyacyl residue. Indeed, a variety of long chain fatty acyl residues arecontemplated including, but not limited to, residues of the followingacids: pentadecanoic acid, palmitic acid, palmitoleic acid,heptadecanoic acid, stearic acid, elaidic acid, oleic acid, nonadecanoicacid, eicosanoic acid, cis-11-eicosenoic acid, 11,14-eicosadienoic acid,heneicosanoic acid, docosanoic acid, erucic acid, tricosanoic acid,tetracosanoic acid, nervonic acid, pentacosanoic acid, hexacosanoicacid, heptacosanoic acid, octacosanoic acid, nonacosanoic acid,triacosanoic acid, vaccenic acid, tariric acid, and ricinoleic acid. Thepresent invention also contemplates powders, oils, food compositions,and pharmaceutical compositions comprising the foregoing acylglycerides.In some embodiments, these compositions further comprise an antioxidant.In some particularly preferred embodiments, the food composition is afunctional food, nutritional supplement food, infant food, pregnancyfood, or elderly food. In other preferred embodiments, the nutritionalor pharmaceutical compositions comprise one of the forgoingacylglycerides and a carrier suitable for oral, intraintestinal, orparenteral administration.

In still other embodiments, the present invention providesacylglycerides having the following structure:

wherein R1 and R3 are acyl residues selected from the group consistingof medium chain and long claim fatty acyl residues and R2 is selectedfrom the group consisting of 10,12; 9,11; 8,10; and 11,13octadecadienoate residues. The present invention is not limited toacylglycerides containing any particular medium chain fatty acylresidue. Indeed, a variety of medium chain fatty acyl residues arecontemplated including, but not limited to, residues of the followingacids: decanoic acid, undecanoic acid, 10-undecanoic acid, lauric acid,cis-5-dodecanoic acid, tridecanoic acid, myristic acid, and myristoleicacid. The present invention is not limited to acylglycerides containingany particular long chain fatty acyl residue. Indeed, a variety of longchain fatty acyl residues are contemplated including, but not limitedto, residues of the following acids: pentadecanoic acid, palmitic acid,palmitoleic acid, heptadecanoic acid, stearic acid, elaidic acid, oleicacid, nonadecanoic acid, eicosanoic acid, cis-11-eicosenoic acid,11,14-eicosadienoic acid, heneicosanoic acid, docosanoic acid, erucicacid, tricosanoic acid, tetracosanoic acid, nervonic acid, pentacosanoicacid, hexacosanoic acid, heptacosanoic acid, octacosanoic acid,nonacosanoic acid, triacosanoic acid, vaccenic acid, tariric acid, andricinoleic acid. The present invention also contemplates powders, oils,food compositions, and pharmaceutical compositions comprising theforegoing acylglycerides. In some embodiments, these compositionsfurther comprise an antioxidant. In some particularly preferredembodiments, the food composition is a functional food, nutritionalsupplement food, infant food, pregnancy food, or elderly food. In otherpreferred embodiments, the nutritional or pharmaceutical compositionscomprise one of the forgoing acylglycerides and a carrier suitable fororal, intraintestinal, or parenteral administration.

In still other embodiments, the present invention providesacylglycerides having the following structure:

wherein R1 and R3 are acyl residues selected from the group consistingof 10,12; 9,11; 8,10; and 11,13 octadecadienoate and R2 is selected fromthe group consisting of ω3, ω6, and ω9 fatty acyl residues. The presentinvention is not limited acylglycerides comprising any particular ω3fatty acyl residue. Indeed, acylglycerides comprising a variety of ω3fatty acyl residues are contemplated, including those selected from thegroup consisting of 9,12,15-octadecatrienoate;6,9,12,15-octadecatetraenoate; 11,14,17-eicosatrienoate;8,11,14,17-eicosatetraenoate; 5,8,11,14,17-eicosapentaenoate;7,10,13,16,19-docosapentaenoate; and 4,7,10,13,16,19-docosahexaenoate.The present invention is not limited to acylglycerides comprising andparticular ω6 fatty acyl residue. Indeed, acylglycerides comprising avariety of ω6 fatty acyl residues are contemplated, including thoseselected from the group consisting of 6,9,12-octadecatrienoate;8,11,14-eicosatrienoate; 5,8,11,14-eicosatetraenoate;7,10,13,16-docosatetraenoate and 4,7,10,13,16-docosapentaenoate. Thepresent invention is not limited to acylglycerides comprising andparticular ω9 fatty acyl residue. Indeed, acylglycerides comprising avariety of ω9 fatty acyl residues are contemplated, including thoseselected from the group consisting of 6,9-octadecadienoate;8,11-eicosadienoate; and 5,8,11-eicosatrienoate. The present inventionalso contemplates powders, oils, food compositions, and pharmaceuticalcompositions comprising the foregoing acylglycerides. In someembodiments, these compositions further comprise an antioxidant. In someparticularly preferred embodiments, the food composition is a functionalfood, nutritional supplement food, infant food, pregnancy food, orelderly food. In other preferred embodiments, the nutritional orpharmaceutical compositions comprise one of the forgoing acylglyceridesand a carrier suitable for oral, intraintestinal, or parenteraladministration.

In still further embodiments, the present invention providesacylglycerides having the following structure:

wherein R1 and R3 are acyl residues selected from the group consistingof ω3, ω6, and ω9 fatty acyl residues and R2 is selected from the groupconsisting 10,12; 9,11; 8,10; and 11,13 octadecadienoate residues. Thepresent invention is not limited acylglycerides comprising anyparticular ω3 fatty acyl residue. Indeed, acylglycerides comprising avariety of ω3 fatty acyl residues are contemplated, including thoseselected from the group consisting of 9,12,15-octadecatrienoate;6,9,12,15-octadecatetraenoate; 11,14,17-eicosatrienoate;8,11,14,17-eicosatetraenoate; 5,8,11,14,17-eicosapentaenoate;7,10,13,16,19-docosapentaenoate; and 4,7,10,13,16,19-docosahexaenoate.The present invention is not limited to acylglycerides comprising andparticular ω6 fatty acyl residue. Indeed, acylglycerides comprising avariety of ω6 fatty acyl residues are contemplated, including thoseselected from the group consisting of 6,9,12-octadecatrienoate;8,11,14-eicosatrienoate; 5,8,11,14-eicosatetraenoate;7,10,13,16-docosatetraenoate and 4,7,10,13,16-docosapentaenoate. Thepresent invention is not limited to acylglycerides comprising andparticular ω9 fatty acyl residue. Indeed, acylglycerides comprising avariety of ω9 fatty acyl residues are contemplated, including thoseselected from the group consisting of 6,9-octadecadienoate;8,11-eicosadienoate; and 5,8,11-eicosatrienoate. The present inventionalso contemplates powders, oils, food compositions, and pharmaceuticalcompositions comprising the foregoing acylglycerides. In someembodiments, these compositions further comprise an antioxidant. In someparticularly preferred embodiments, the food composition is a functionalfood, nutritional supplement food, infant food, pregnancy food, orelderly food. In other preferred embodiments, the nutritional orpharmaceutical compositions comprise one of the forgoing acylglyceridesand a carrier suitable for oral, intraintestinal, or parenteraladministration.

In still other embodiments, the present invention providesacylglycerides having the following structure:

wherein R1 and R3 are 10,12 octadecadienoate and R2 is 9,11octadecadienoate. In some preferred embodiments, the 10,12octadecadienoate is t10,c12 octadecadienoate and the 9,11octadecadienoate is c9,t11 octadecadienoate.

In some embodiments, the present invention provides acylglycerideshaving the following structure:

wherein R1 and R3 are 9,11 octadecadienoate and R2 is 10,12octadecadienoate. In some preferred embodiments, the 10,12octadecadienoate is t10,c12 octadecadienoate and the 9,11octadecadienoate is c9,t11 octadecadienoate.Definitions

As used herein, “conjugated linoleic acid” or “CLA” refers to anyconjugated linoleic acid or octadecadienoic free fatty acid. It isintended that this term encompass and indicate all positional andgeometric isomers of linoleic acid with two conjugated carbon-carbondouble bonds any place in the molecule. CLA differs from ordinarylinoleic acid in that ordinary linoleic acid has double bonds at carbonatoms 9 and 12. Examples of CLA include cis- and trans isomers (“E/Zisomers”) of the following positional isomers: 2,4-octadecadienoic acid,4,6-octadecadienoic acid, 6,8-octadecadienoic acid, 7,9-octadecadienoicacid, 8,10-octadecadienoic acid, 9,11-octadecadienoic acid and 10,12octadecadienoic acid, 11, 13 octadecadienoic acid. As used herein, “CLA”encompasses a single isomer, a selected mixture of two or more isomers,and a non-selected mixture of isomers obtained from natural sources, aswell as synthetic and semisynthetic CLA.

As used herein, the term “isomerized conjugated linoleic acid” refers toCLA synthesized by chemical methods (e.g., aqueous alkali isomerization,non-aqueous alkali isomerization, or alkali alcoholate isomerization).

As used herein, the term “conjugated linoleic acid moiety” refers to anycompound or plurality of compounds containing conjugated linoleic acidsor derivatives. Examples include, but are not limited to fatty acids,alkyl esters, and triglycerides of conjugated linoleic acid.

As used herein, it is intended that “triglycerides” or “acylglycerides”of CLA contain CLA at any or all of three positions (e.g., SN-1, SN-2,or SN-3 positions) on the triglyceride backbone. Accordingly, atriglyceride containing CLA may contain any of the positional andgeometric isomers of CLA.

As used herein, it is intended that “esters” of CLA include any and allpositional and geometric isomers of CLA bound through an ester linkageto an alcohol or any other chemical group, including, but not limited tophysiologically acceptable, naturally occurring alcohols (e.g.,methanol, ethanol, propanol). Therefore, an ester of CLA or esterifiedCLA may contain any of the positional and geometric isomers of CLA.

It is intended that “non-naturally occurring isomers” of CLA include,but are not limited to c11,t13; t11,c13; t11,t13; c11,c13; c8,t10;t8,c10; t8,t10; c8,c10; and trans-trans isomers of octadecadienoic acid,and does not include t10,c12 and c9,t11 isomers of octadecadienoic acid.“Non-naturally occurring isomers” may also be referred to as “minorisomers” of CLA as these isomers are generally produced in low amountswhen CLA is synthesized by alkali isomerization.

As used herein, “low impurity” CLA refers to CLA compositions, includingfree fatty acids, alkylesters, and triglycerides, which contain lessthan 1% total 8,10 octadecadienoic acids, 11,13 octadecadienoic acids,and trans-trans octadecadienoic acids. As used herein, “c” encompasses achemical bond in the cis orientation, and “t” refers to a chemical bondin the trans orientation. If a positional isomer of CLA is designatedwithout a “c” or a “t”, then that designation includes all four possibleisomers. For example, 10,12 octadecadienoic acid encompasses c10,t12;t10,c12; t10,t12; and c10,c12 octadecadienoic acid, while t10,c12octadecadienoic acid or CLA refers to just the single isomer.

As used herein, the term “oil” refers to a free flowing liquidcontaining long chain fatty acids (e.g., CLA), triglycerides, or otherlong chain hydrocarbon groups. The long chain fatty acids, include, butare not limited to the various isomers of CLA.

As used herein, the term “physiologically acceptable carrier” refers toany carrier or excipient commonly used with oily pharmaceuticals. Suchcarriers or excipients include, but are not limited to, oils, starch,sucrose and lactose.

As used herein, the term “oral delivery vehicle” refers to any means ofdelivering a pharmaceutical orally, including, but not limited to,capsules, pills, tablets and syrups.

As used herein, the term “food product” refers to any food or feedsuitable for consumption by humans, non-ruminant animals, or ruminantanimals. The “food product” may be a prepared and packaged food (e.g.,mayonnaise, salad dressing, bread, or cheese food) or an animal feed(e.g., extruded and pelleted animal feed or coarse mixed feed).“Prepared food product” means any pre-packaged food approved for humanconsumption.

As used herein, the term “foodstuff” refers to any substance fit forhuman or animal consumption.

As used herein, the term “functional food” refers to a food product towhich a biologically active supplement has been added.

As used herein, the term “infant food” refers to a food productformulated for an infant such as formula.

As used herein, the term “elderly food” refers to a food productformulated for persons of advanced age.

As used herein, the term “pregnancy food” refers to a food productformulated for pregnant women.

As used herein, the term “nutritional supplement” refers to a foodproduct formulated as a dietary or nutritional supplement to be used aspart of a diet.

As used herein, the term “medium chain fatty acyl residue” refers tofatty acyl residues derived from fatty acids with a carbon chain lengthof equal to or less than 14 carbons.

As used herein, the term “long chain fatty acyl residue” refers to fattyacyl residues derived from fatty acids with a carbon chain length ofgreater than 14 carbons.

As used herein, the term “volatile organic compound” refers to any smallcarbon-containing compound which exists partially or completely in agaseous state at a given temperature. Volatile organic compounds may beformed from the oxidation of an organic compound (e.g., CLA). Volatileorganic compounds include, but are not limited to pentane, hexane,heptane, 2-butenal, ethanol, 3-methyl butanal, 4-methyl pentanone,hexanal, heptanal, 2-pentyl furan, octanal.

As used herein, the term “metal oxidant chelator” refers to anyantioxidant that chelates metals. Examples include, but are not limitedto lecithin and citric acid esters.

As used herein, the term “alcoholate catalyst” refers to alkali metalcompounds of any monohydric alcohol, including, but not limited to,potassium methylate and potassium ethylate.

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to the field of human and animalnutrition, and in particular to certain novel compositions of conjugatedlinoleic acids (CLA). In particular, the present invention relates tocertain isomers of conjugated linoleic acids at either the SN1 and SN3or SN2 positions of the acylglyceride molecule and another fatty acylresidue at the other of the SN1 and SN3 or SN2 positions of theacylglyceride molecule.

I. Sources of Conjugated Linoleic Acids

The acylglycerides of the present invention contain conjugated linoleicacyl residues. The conjugated linoleic acid incorporated in theseacylglycerides may be made by a variety of methods, for example, thosedescribed in U.S. Pat. Nos. 6,015,833 and 6,060,514, each of which isherein incorporated by reference. In some embodiments, sunflower oil,safflower oil, or corn oil are reacted at an ambient pressure under aninert gas atmosphere with an excess of alkali in a high-boiling pointsolvent, namely propylene glycol at a temperature below the boilingpoint of the solvent. In some particularly preferred embodiments,sunflower oil, safflower oil, or corn oil are reacted in the presence ofan alkali alcoholate catalyst and a small amount of a suitable solvent.As compared to soybean oil, these oils have lower concentrations ofundesirable components such as phosphatides and sterols. Theseundesirable components may contribute to the formation of gums whichfoul the conjugation equipment and other undesirable polymers.

A. Isomerization with Propylene Glycol as a Solvent

In some embodiments of the present invention, the conjugated linoleicacid is produced by nonaqueous alkali isomerization. The reactionconditions of the controlled isomerization process allow for precisecontrol of the temperature (and constant ambient pressure) of theconjugation process. Preferably the alkali is an inorganic alkali suchas potassium hydroxide, cesium hydroxide, cesium carbonate or an organicalkali such as tetraethyl ammonium hydroxide. The catalyst is preferablyprovided in a molar excess as compared to the fatty acid content of oil.The solvent is propylene glycol. Preferably, the reaction is conductedwithin a temperature range 130 to 165° C., most preferably at about 150°C. The time of the reaction may vary, however, there is an increasedlikelihood of the formation of undesirable isomers when the reaction isconducted for long periods of time. A relatively short reaction time of2.0 to 6.5 hours has proved satisfactory for excellent yields.

It will be understood to a person skilled in the art that to produce thedesired composition, the reaction conditions described above may bevaried depending upon the oil to be conjugated, the source of alkali,and equipment. Preanalysis of a particular oil may indicate that theconditions must be varied to obtain the desired composition. Therefore,the temperature range, pressure, and other reaction parameters representa starting point for design of the individual process and are intendedas a guide only. For example, it is not implied that the describedtemperature range is the only range which may be used. The essentialaspect is to provide precise temperature control. However, care must betaken because increasing the pressure may lead to less than completeisomerization and the formation of undesirable isomers. Finally, thelength of the conjugation reaction may be varied. Generally, increasingamounts of undesirable isomers are formed with increasing length orreaction time. Therefore, the optimal reaction time allows the reactionto go nearly or essentially to completion but does not result in theformation of undesirable isomers.

Following the conjugation reaction, the resulting CLA containingcomposition may be further purified. To separate the fatty acids fromthe conjugation reaction mix, the reaction mix is cooled toapproximately 95° C., an excess of water at 50° C. is added, and themixture slowly stirred while the temperature is reduced to about 50° C.to 60° C. Upon addition of the water, a soap of the fatty acids isformed and glycerol is formed as a by-product. Next, a molar excess ofconcentrated HCl is added while stirring. The aqueous and nonaqueouslayers are then allowed to separate at about 80-90° C. The bottom layercontaining water and propylene glycol is then drawn off. The remainingpropylene glycol is removed by vacuum dehydration at 60-80° C.

The dried CLA composition may then preferably be degassed in degassingunit with a cold trap to remove any residual propylene glycol. Next, theCLA is distilled at 190° C. in a molecular distillation plant at avacuum of 10⁻¹ to 10⁻² millibar. The advantage of this purificationsystem is the short time (less than one minute) at which the CLA is heldat an elevated temperature. Conventional batch distillation proceduresare to be strictly avoided since they involve an elevated temperature ofapproximately 180-200° C. for up to several hours. At these elevatedtemperatures the formation of undesirable trans-trans isomers willoccur. Approximately 90% of the feed material is recovered as a slightlyyellow distillate. The CLA may then be deodorized by heating to about120°-170° C., preferably at about 150° C. for 2 hours to improve smelland taste. Excessive heat may result in the formation of trans-transisomers. These procedures produce a CLA composition with a solvent levelof less than about 5 ppm, preferably less than about 1 ppm. This processeliminates toxic trace levels of solvent so that the resultingcomposition is essentially free of toxic solvent residues.

The processes described above are readily adaptable to both pilot andcommercial scales. For example, 400 kg of safflower oil may beconjugated at 150° C. for 5 hours in 400 kg of propylene glycol with 200kg KOH added as a catalyst. The resulting CLA may then be purified asdescribed above. Further, commercial scale batch systems may be easilymodified to produce the desired CLA composition. For example, stainlesssteel reactors should be preferably glass lined to prevent corrosion dueto pH levels of below 3.0. However, it should be noted that conjugationprocesses utilizing nonaqueous solvents are generally less corrosivethan those conducted with water.

B. Isomerization with Alcoholate Catalysts

In other embodiments, the acylglycerides of the present inventionincorporate acylglycerides made by the isomerization of linoleic acid inthe presence alcoholate catalysts. After fat splitting and dehydration,the free fatty acids are combined with methanol or another monohydriclow molecular weight alcohol and heated to the temperature at which thealcohol boils. Esterification proceeds under refluxing conditions withremoval of the reaction water through a condenser. After the addition ofa further quantity of the same or a different monohydric alcohol analcoholate catalyst is blended into the ester mix (See, e.g., U.S. Pat.No. 3,162,658, incorporated herein by reference). Typical alcoholatecatalysts are sodium or potassium ethoxide, or their methyl, butyl, orpropyl counterparts.

In the esterification, methanol or ethanol are preferred, although otherbranched or straight chain monohydric alcohols may be used. The longerthe aliphatic chain of the alkyl group, the more lipid compatible thematerial becomes. Also the viscosity tends to increase. For differenttypes of feed or food, whose consistency varies, product of varyingviscosity can be used to obtain the desired flow or compoundingcharacteristics without affecting the therapeutic or nutritionalproperties arising from the CLA moieties. The theory and practice ofesterification are conventional. A basic explanation of the most commonmethods is set forth in the McCraw-Hill Encyclopedia of Science &Technology, McGraw-Hill Book Co., N.Y.: 1996 (5th ed.). The animal andhuman body has a variety of esterases, so that the CLA-ester is cleavedto release the free fatty acids readily. Tissue uptake may have adifferent kinetics depending on the tissue involved and the benefitsought.

In the isomerization step, it was found that alcoholate catalysisproduced a much superior product than aqueous alkali mediatedisomerization. The latter process always produced undesirable isomerseven under mild reaction conditions. The milder conditions do give loweramounts of unwanted isomers, but at the great expense of yield, as shownin the Examples. In most systems the appearance of the c9,t11 andt10,c12 isomers dominates and they are formed in roughly equimolaramounts. It has not heretofore been possible to control theisomerization of the one isomer to the exclusion of the other. While itis desirable to increase the percentage of one or the other isomer(depending on the physiological effect to be achieved), at present thismust largely be carried out by adding an enriched source of the desiredisomer.

The preferred starting materials for conjugation with alcoholatecatalysts are sunflower oil, safflower oil, and corn oil. Each of theseoils contains high levels of linoleic acid and low levels of linolenicacid. Conjugation of linolenic acid results in the formation of severaluncharacterized fatty acid moieties, the biological properties of whichare unknown. Previous conjugation processes were not concerned with theproduction of unknown compounds because the products were used in dryingoils, paints and varnishes and not in products destined from human oranimal consumption.

In some embodiments, it is further contemplated that glycerol and estersof glycerol should be removed before making monoesters of fatty acids.Traces of glycerol present during conjugation contribute to theproduction of trimethoxypropane and triethoxypropane. Therefore, priorto conjugation, it is preferable to distill monoesters obtained byalcoholysis.

C. Synthesis of Other CLA Isomers

The present invention also contemplates the synthesis of triglyceridescomprising the isomers listed in Table 1 below. In some embodiments ofthe invention, a partially purified or concentrated isomer of CLA istreated under conditions that cause migration of the double bond system.In preferred embodiments, the conditions comprise heating at least oneisomer to about 200-240° C., preferably to about 220° C. In otherembodiments, the conditions further comprise reacting the partiallypurified or concentrated isomer or isomers under nitrogen in a sealedcontainer. Referring to Table 1, the preparations of isomers in column 1can be used to produce preparations containing a substantial amount ofthe corresponding isomer in column 2. After the initial conversionreaction, the preparation will contain both the starting isomer and the“sister” isomer. Likewise, the preparations of isomers in column 2 canbe used to produce substantial amounts of the corresponding isomer incolumn 1. The preparations containing both isomers may be furthertreated to purify the sister isomer (e.g., by gas chromatography). Aswill be understood by those skilled in the art, it is possible to startwith more than one partially purified isomer, thereby producing apreparation containing four, six, eight or more isomers. In furtherembodiments, a purified preparation of the sister isomer may be preparedby methods known in the art (i.e., gas-liquid chromatography) from thetreated preparation containing the initial isomer and its sister isomer.

TABLE 1 Column 1 Column 2 c9, t11 t8, c10 t10, c12 c11, t13 c7, t9 t6,c8 t11, c13 c12, t14 c6, t8 t5, c6 c5, t7 t4, c6 c4, t6 t3, c5 t12, c14c13, t15 t13, c15 c14, t16

As demonstrated in the Examples, treatment of purified t10,c12octadecadienoic acid resulted in the production of c11,t13octadecadienoic acid. Likewise, concentrated or partially purifiedc11,t13 octadecadienoic acid can be used to produce t10,c12octadecadienoic acid.

D. Other Sources of Conjugated Linoleic Acid Isomers

In other embodiments, the conjugated linoleic acids used to produce theacylglycerides of the present invention are obtained from alternativesources. For example, some isomers (e.g., t10,c12 and c9,t11) areavailable from commercial sources. In other embodiments, t10,c12 andc9,t11 CLA may be purified by the methods described in Scholfield etal., JAOCS 47(8):303 (1970) and Berdeau et al., JAOCS 74:1749-55 (1998).This method allows for the crystallization and precipitation of thet10,c12 isomer from a mixture of isomers. If the initial mixturecontains predominantly the t10,c12 and c9,t11 isomers (i.e., theisomerization id conducted as described above), then the oil remainingafter precipitation will be enriched for c9,t11 CLA. In still furtherembodiments, the CLA isomers may be prepared by gas chromatography orgas chromatography/mass spectrometry procedures.

II. Synthesis of Triglycerides

The present invention provides novel acylglycerides, as well as foodcompositions, animal feeds, pharmaceutical compositions and nutritionalcompositions comprising the novel acylglycerides. According to thepresent invention acylglycerides are provided having the followinggeneral structure:

wherein R1 and R3 are acyl residues selected from the group consistingof 10,12; 9,11; 8,10; and 11,13 octadecadienoate and R2 is selected fromthe group consisting of long chain and medium chain fatty acyl residues.

In other embodiments, acylglycerides having the following generalstructure are provided:

wherein R1 and R3 are acyl residues selected from the group consistingof medium chain and long claim fatty acyl residues and R2 is selectedfrom the group consisting of 10,12; 9,11; 8,10; and 11,13octadecadienoate residues.

In other embodiments, acylglycerides having the following generalstructure are provided:

wherein R1 and R3 are acyl residues selected from the group consistingof 10,12; 9,11; 8,10; and 11,13 octadecadienoate and R2 is selected fromthe group consisting of ω3, ω6, and ω9 fatty acyl residues.

In other embodiments, acylglycerides having the following generalstructure are provided:

wherein R1 and R3 are acyl residues selected from the group consistingof ω3, ω6, and ω9 fatty acyl residues and R2 is selected from the groupconsisting 10,12; 9,11; 8,10; and 11,13 octadecadienoate residues.

In other embodiments, acylglycerides having the following generalstructure are provided:

wherein R1 and R3 are 10,12 octadecadienoate and R2 is 9,11octadecadienoate.

In other embodiments, acylglycerides having the following generalstructure are provided:

wherein R1 and R3 are 9,11 octadecadienoate and R2 is 10,12octadecadienoate.

The present invention is not limited to acylglycerides comprisingresidues of any particular isomer of conjugated linoleic acid. Indeed,the use of a variety of isomers of conjugated linoleic acid iscontemplated, including, but not limited to t10,c12 octadecadienoate;c10,t12 octadecadienoate; c9,t11 octadecadienoate; t9,c11octadecadienoate; c8,t10 octadecadienoate; t8,c10 octadecadienoate;t11,c13 octadecadienoate; and c11,t13 octadecadienoate, as well as theother isomers listed in Table 1 above.

The present invention is not limited to acylglycerides comprising anyparticular long chain or medium chain fatty acid residues. Indeed, theincorporation of a variety long chain and medium chain fatty acidresidues is contemplated, including, but not limited to decanoic acid(10:0), undecanoic acid (11:0), 10-undecanoic acid (11:1), lauric acid(12:0), cis-5-dodecanoic acid (12:1), tridecanoic acid (13:0), myristicacid (14:0), myristoleic acid (cis-9-tetradecenoic acid, 14:1),pentadecanoic acid (15:0), palmitic acid (16:0), palmitoleic acid(cis-9-hexadecenoic acid, 16:1), heptadecanoic acid (17:1), stearic acid(18:0), elaidic acid (trans-9-octadecenoic acid, 18:1), oleic acid(cis-9-octadecanoic acid, 18:1), nonadecanoic acid (19:0), eicosanoicacid (20:0), cis-11-eicosenoic acid (20:1), 11,14-eicosadienoic acid(20:2), heneicosanoic acid (21:0), docosanoic acid (22:0), erucic acid(cis-13-docosenoic acid, 22:1), tricosanoic acid (23:0), tetracosanoicacid (24:0), nervonic acid (24:1), pentacosanoic acid (25:0),hexacosanoic acid (26:0), heptacosanoic acid (27:0), octacosanoic acid(28:0), nonacosanoic acid (29:0), triacosanoic acid (30:0), vaccenicacid (t-11-octadenecoic acid, 18:1), tariric acid (octadec-6-ynoic acid,18:1), and ricinoleic acid (12-hydroxyoctadec-cis-9-enoic acid, 18:1).

The present invention is not limited to acylglycerides comprising anyparticular ω3, ω6, and ω9 fatty acyl residues. Indeed, the presentinvention encompasses, but is not limited to, acylglycerides includingresidues of the following ω3, ω6, and ω9 fatty acids:

-   9,12,15-octadecatrienoic acid (α-linolenic acid) [18:3, ω3];-   6,9,12,15-octadecatetraenoic acid (stearidonic acid) [18:4, ω3];-   11,14,17-eicosatrienoic acid (dihomo-α-linolenic acid) [20:3, ω3];-   8,11,14,17-eicosatetraenoic acid [20:4, ω3],-   5,8,11,14,17-eicosapentaenoic acid [20:5, ω3];-   7,10,13,16,19-docosapentaenoic acid [22:5, ω3];-   4,7,10,13,16,19-docosahexaenoic acid [22:6, ω3];-   9,12-octadecadienoic acid (linoleic acid) [18:2, ω6];-   6,9,12-octadecatrienoic acid (γ-linolenic acid) [18:3, ω6];-   8,11,14-eicosatrienoic acid (dihomo-γ-linolenic acid) [20:3 ω6];-   5,8,11,14-eicosatetraenoic acid (arachidonic acid) [20:4, ω6];-   7,10,13,16-docosatetraenoic acid [22:4, ω6];-   4,7,10,13,16-docosapentaenoic acid [22:5, ω6];-   6,9-octadecadienoic acid [18:2, ω9];-   8,11-eicosadienoic acid [20:2, ω9]; and-   5,8,11-eicosatrienoic acid (Mead acid) [20:3, ω9].

Moreover, acyl residues may be hydroxylated, epoxidated orhydroxyepoxidated acyl residues.

In some embodiments of the present invention, the novel acylglyceridesof the present invention are prepared by chemical method synthesis.These methods and interemediates for use therein are described inexamples 8-20.

In other embodiments, novel acylglycerides of the present invention aremanufactured by using non-specific and position-specific lipases toinsert a first fatty acyl residue at position 2 (SN2) of theacylglyceride and a second fatty acyl residue at positions 1 and 3 (SN1and SN3) of the acylgyceride. Non-specific lipases are lipases that areable to hydrolyse or esterify (i.e., the reverse reaction) fatty acidsin all positions on a glycerol. A position-specific or 1,3 specificlipase almost exclusively hydrolyses or esterifies fatty acids inposition 1 and 3 on the glycerol backbone. The structured acylglyceridesof the present invention are synthesized by first using a non-specifclipase to attach the desired fatty acid for position 2 to all 3positions and then hydrolysing the acyl residues in position 1 and 3using a 1,3 specific lipase. The hydrolysed acids are then removed bydistillation before the acids desired to be attached to positions 1 are3 are added and esterified to position 1 and 3 by the same lipase. Thedirection of the reaction (hydrolysis or esterification) is easilycontrolled by water addition or removal respectively. In the followingexample is a general outline of the method.

In particularly preferred embodiments, a purified aliquot of a firstfatty acid (about 3 moles), glycerol (about 1 mole) and up to 10% byweight of acids are mixed with immobilized non-specific lipase(commercially available). The mixture is stirred under vacuum andslightly heated (50-60° C.). The water produced during theesterification is continuously removed by the vacuum suction. After24-48 hours, the reaction is finished and the enzymes are removed andrecovered by filtration. The resulting acylglyceride has the first fattyacid attached at all three positions. The first fatty acid residue atpositions 1 and 3 is then removed in by addition of 1,3 specificimmobilized lipase (commercially available) and 1% water. The mixture isheated to 50-60° C. and stirred under nitrogen atmosphere for 24-48hours. The reaction mixture now comprises free fatty acids liberatedfrom position 1 and 3 and monoglycerides (fatty acid B attached toposition 2). Next, in preferred embodiments, the fatty acids aredistilled off from the mixture by molecular distillation. In furtherpreferred embodiments, about one mole of the monoglyceride is allowed toreact for 24-48 hours with 2 moles a second free fatty acid in thepresence of 1,3 specific lipase. In some embodiments, this reactiontakes place under stirring and vacuum at 50-60° C. to remove waterproduced in the esterification process. The resulting acylglyceride is astructured triglyceride with the first fatty acid in position 2 and thesecond fatty acid in positions 1 and 3.

As described above, in some embodiments of the present invention, lipasethat specifically acts on the positions 1 and 3 of triglyceride is usedas catalyst. The present invention is not limited to the use of anyparticular 1,3 specific lipase. Examples of 1,3 specific lipases usefulin the present invention include lipases produced by a microorganismbelonging to the genus Rhizopus, Rhizomucor, Mucor, Penicillium,Aspergillus, Humicola or Fusarium, as well as porcine pancreatic lipase.Examples of commercially available lipases include lipase of Rhizopusdelemar (Tanabe Pharmaceutical, Dalipase), lipase of Rhizomucor miehei(Novo Nordisk, Ribozyme IM), lipase of Aspergillus niger (AmanoPharmaceutical, Lipase A), lipase of Humicola lanuginosa (Novo Nordisk,Lipolase), lipase of Mucor javanicus (Amano Pharmaceutical, Lipase M)and lipase of Fusarium heterosporum. These lipases may be used in theirnative form, or in the form of lipase that has been immobilized oncellite, ion exchange resin or a ceramic carrier.

The amount of water added to the reaction system affects the outcome ofthe reaction. Transesterification does not proceed in the absoluteabsence of water, while if the amount of water is too much, hydrolysisoccurs, the triglyceride recovery rate decreases, or spontaneous acylgroup transfer occurs in a partially acylated glyceride resulting intransfer of the saturated fatty acid at the position 2 to the position 1or 3. Thus, when using an immobilized enzyme that does not have bondedwater, it is effective to first activate the enzyme using a substrate towhich water has been added before carrying out the reaction, and thenuse a substrate to which water is not added during the reaction. Inorder to activate the enzyme in batch reactions, a substrate containingwater at 0 to 1,000% (wt %) of the amount of added enzyme should be usedto pretreat the enzyme, and in the case of activating by a columnmethod, a water-saturated substrate should be allowed to continuouslyflow through the column. The amount of lipase used in a batch reactionmay be determined according to the reaction conditions. Although thereare no particular limitations on the amount of lipase, 1 to 30% (wt %)of the reaction mixture is suitable when using, for example, lipase ofRhizopus delemar or lipase of Rhizomucor miehei immobilized on celliteor a ceramic carrier.

In some preferred embodiments, the above-mentioned immobilized enzymecan be used repeatedly. Namely, the reaction can be continued by leavingthe immobilized enzyme in a reaction vessel after reaction and replacingthe reaction mixture with freshly prepared reaction mixture comprisingsubstrate. In addition, for transesterification by a column method, areaction mixture containing substrate be allowed to flow continuously atthe rate of 0.05 to 20 ml/hr per gram of enzyme. In other preferredembodiments, the content of target triglyceride can be increased byperforming transesterification repeatedly. Namely, lipase specificallyacting on the positions 1 and 3 of the acylglyceride is allowed to actin the presence of the second fatty acid or an ester thereof to obtain areaction mixture in which fatty acids at positions 1 and 3 aretransesterified to the desired fatty acid.

The target acylglycerides of the present invention can easily beisolated by routine methods such as liquid chromatography, moleculardistillation, downstream membrane fractionation or vacuumsuperfractionation or a combination thereof. Purification of the targetacylgycerides of the present invention can be performed by alkalinedeacidation, steam distillation, molecular distillation, downstreammembrane fractionation, vacuum superfractionation, columnchromatography, solvent extraction or membrane separation, or acombination thereof so as to remove the above-mentioned fatty acidsreleased by the transesterification and unreacted unsaturated fattyacids.

III. Stabilization of CLA Acylglycerides

The present invention also contemplates stabilization of the CLAacylglycerides by preventing oxidation of the compounds. The presentinvention is not limited to any one mechanism. Indeed, an understandingof the mechanism of the invention is not necessary to produce thecomposition or perform the methods of the present invention.Nevertheless, unlike non-conjugated fatty acids, CLA does not appear toform peroxide breakdown products. This was demonstrated experimentallyby measuring peroxide values (PV) spectrophotometrically by achlorimetirc ferric thiocyanate method. After storage in open glass, thePV of CLA was 32; in comparison, the value for linoleic acid was 370.

CLA forms volatile organic compounds during breakdown, including hexane.Products stored in a steel drum for several weeks were found to containup to 25 ppm hexane. Hexane has a characteristic taste and smell that isundesirable in food products. Oxidation of CLA appears to be caused bythe presence of metal contaminants. Thus, a system for removal of suchcompounds that promote oxidation during purification is advantageous.

Furthermore, it is also advantageous to add compounds to CLAacylglycerides to decrease oxidation during storage. Compounds thatprevent oxidation (antioxidants) have two general mechanisms of action.The first is the prevention of oxidation by lipid peroxide radicalscavenging. Examples include but are not limited to tocopherols andascorbylpalmitate. The second mechanism for preventing oxidation is bythe chelation of metal ions. Examples of metal oxidant chelatorsinclude, but are not limited to, citric acid esters and lecithin. Somecommercially available compounds (e.g., Controx, Grumau (Henkel),Illertissen, Del.) include both peroxide scavengers and metal chelators(e.g., lecithin, tocopherols, ascorbylpalmitate, and citric acidesters). In some embodiment of the present invention, metal oxidantchelators are added to CLA containing compounds to prevent oxidation. Inother embodiments, a combination of metal oxidant chelators and peroxidescavengers is included in the CLA acylglyceride composition.

In some embodiments, gas chromatography/mass spectroscopy is used indetect the presence of volatile organic breakdown products of CLA. Inother embodiments, oil stability index (OSI) measurements are used todetect the presence of volatile organic breakdown products of CLA. Insome embodiments of the present invention, pro-oxidants (e.g., iron) areremoved from the CLA acylglyceride compositions. Methods for removingpro-oxidants include, but are not limited to, distillation or byadsorption. In some embodiments of the present invention, compounds areadded to prevent oxidation of CLA.

In preferred embodiments, precautions are taken during purification toprevent oxidation during storage. These precautions include the removalof compounds that serve as pro-oxidants, including but not limited toiron or other metals. In some embodiments, metals are removed bytreating with adsorbing agents, including but not limited to bleachingearth, active charcoal zeolites, and silica. In other embodiments, thepro-oxidants are removed by distillation.

In some embodiments, pro-oxidants are removed in a distillation process.In some preferred embodiments, distillation of a CLA acylglyceride ofthe present invention is performed on a molecular distillationapparatus. Distillation is carried out at 150° C. and a pressure of 10⁻²mbar. The present invention is not intended to be limited to theconditions described for distillation. Other temperatures and pressuresare within the scope of the present invention.

In some embodiments, oxidation of the CLA acylglycerides of the presentinvention is prevented by the addition of metal oxidant chelators orperoxide scavengers to the finished product. In some embodiments, theamount of oxidation is measured by the oil stability index (OSI). TheOSI (See e.g., AOCS official method Cd 12b-92) is a measurement of anoil's resistance to oxidation. It is defined mathematically as the timeof maximum change of the rate of oxidation. This rate can be determinedmathematically. Experimentally, the OSI is calculated by measuring thechange in conductivity of deionized water is which volatile organicacids (oxidation products) are dissolved. When performing OSImeasurements, it is important to avoid contamination by trace amounts ofmetals, which can accelerate the oxidation process. This is generallyaccomplished by careful washing of all glassware used with a cleaningsolution lacking chromate or surfactants. Water must be deionized andall solvents must be of a highly purified grade.

IV. Formulation and Administration of CLA Acylglycerides

The CLA acylglycerides of the present invention may be provided in avariety of forms. In some embodiments, administration is oral. The CLAmoieties may be formulated with suitable carriers such as starch,sucrose or lactose in tablets, pills, dragees, capsules, solutions,liquids, slurries, suspensions and emulsions. Preferably, the CLAformulations contain antioxidants, including, but not limited toControx, Covi-OX, lecithin, and oil soluble forms of vitamin C (ascorbylpalmitate). The CLA acylglyceride may be provided in oily solution, orin any of the other forms discussed above. The tablet or capsule of thepresent invention may be coated with an enteric coating which dissolvesat a pH of about 6.0 to 7.0. A suitable enteric coating which dissolvesin the small intestine but not in the stomach is cellulose acetatephthalate. In some embodiments, the CLA is provided as soft gelatincapsules containing 500-1500 mg of acylglyceride. The CLA may also beprovided by any of a number of other routes, including, but not limitedto, intravenous, intramuscular, intra-arterial, intramedullary,intrathecal, intraventricular, transdermal, subcutaneous,intraperitoneal, intranasal, enteral, topical, sublingual or rectalmeans. Further details on techniques for formulation for andadministration and administration may be found in the latest edition ofRemington's Pharmaceutical Sciences (Maack Publishing Co., Easton, Pa.).

In particularly preferred embodiments, the acylglycerides of the presentinvention are combined with an excipient or powdering agent. The mixtureis then formed into a powder by methods such as spray drying (See, e.g.,U.S. Pat. No. 4,232,052, incorporated herein by reference). In general,spray drying involves liquefying or emulsifying a substance and thenatomizing it so that all but a small percentage of water is removed,yielding a free flowing powder. Suitable spray drying units include bothhigh pressure nozzle spray driers and spinning disk or centrifugal spraydriers. The present inventors have discovered that powders containinghigh loads (e.g., 40%-65%) conjugated linoleic acid and/or other oils(e.g., evening primrose oil, borage oil, flax oil, CLA oil) can beformed by the simple spray drying of the emulsion of the oil, excipientand water. It is not necessary to incorporate more complex methodsinvolving spraying into a fluidized bed or spraying in a countercurrentfashion.

The present invention is not limited to any particular excipient.Indeed, a variety of excipients are contemplated, including, but notlimited to, HI-CAP 100 (National Starch, Bridgewater, N.J.) and HI-CAP200 (National Starch, Bridgewater, N.J.). The powder of the presentinvention contains a high percentage of oil as compared to theexcipient. In some embodiments, the oil is 20% of the powder on aweight/weight basis (i.e., the powder contains 20 grams of oil for every100 grams of powder). In other embodiments, the oil is 35% of the powderon weight/weight basis. In still other embodiments, the oil is at least50% of the powder on a weight/weight basis. In further embodiments, theoil is at least 60%-65% of the powder on a weight/weight basis. In eachcase, the oil powder is free flowing and odorless. In preferredembodiments, the oil comprises a CLA moiety. In particularly preferredembodiments, the oil comprises CLA acylglyceries of the presentinvention.

An effective amount of CLA moiety may also be provided as a supplementin various food products, including animal feeds, human functional foodproducts, infant food products, nutritional supplements, and drinks. Forthe purposes of this application, food products containing CLAacylglycerides means any natural, processed, diet or non-diet foodproduct to which exogenous CLA acylglyceride has been added. Therefore,CLA acylglycerides may be directly incorporated into various preparedfood products, including, but not limited to diet drinks, diet bars,supplements, prepared frozen meals, candy, snack products (e.g., chips),prepared meat products, milk, cheese, yogurt and any other fat or oilcontaining foods.

Furthermore, as shown above and in the Examples, CLA acylglyceridecompositions can contain levels of volatile organic compounds that causethe taste and smell of food products containing the CLA acylglyceride tobe adversely effected. It is contemplated that the food products of thepresent invention that contain CLA acylglyceride compositions havingless than 100 ppm volatile organic compounds, and preferably less than 5ppm volatile organic compounds, are superior in taste and smell to foodproducts containing higher levels of volatile organic compounds and willbe preferred in blind taste and smell tests. Accordingly, someembodiments of the present invention provide a food product containing aCLA acylglyceride, wherein the conjugated linoleic acid moiety has asufficiently low volatile organic acid compound concentration so thattaste and smell of the food product is not affected.

Experimental

The following examples are provided in order to demonstrate and furtherillustrate certain preferred embodiments and aspects of the presentinvention and are not to be construed as limiting the scope thereof.

In the experimental disclosure which follows, the followingabbreviations apply: M (molar); mM (millimolar); μM (micromolar); kg(kilograms); g (grams); mg (milligrams); μg (micrograms); ng(nanograms); L or l (liters); ml (milliliters); μl (microliters); cm(centimeters); mm (millimeters); nm (nanometers); ° C. (degreescentigrade); KOH (potassium hydroxide); HCL (hydrochloric acid); Hg(mercury).

EXAMPLE 1

Isomerization of Safflower Oil Using Propylene Glycol at Low Temperature

Safflower oil was isomerized in propylene glycol at low temperaturesusing KOH as a catalyst. The isomerization apparatus consisted of atwo-necked flask with a thermometer placed in one neck, leaving a smallopening to release excess pressure. A nitrogen supply was attached tothe other neck of the flask. Solutions added to the flask were agitatedby the use of a magnetic bar and a magnetic stirrer. The temperature ofthe flask was controlled by placing the flask in a thermostat-controlledoil bath placed on the magnetic stirrer.

The flask was filled with 60.27 g propylene glycol and 28.20 g KOH andimmersed into the oil bath. The temperature was increased to 130° C. todissolve the KOH. After the KOH had dissolved, 60.09 g of safflower oilwas introduced into the flask. A high volume of nitrogen was circulatedthrough the two-neck flask for 5 min. and then reduced to a lowervolume. The mixture was heated to 150° C., which took approximately 40min. The mixture was then allowed to react at 150° C. for 3.5 hours. Atintervals, 3 ml samples were withdrawn for analysis.

The samples were placed directly into 6 ml of hot water and citric acidwas added in excess until the free fatty acids separated out as the toplayer. Heating was necessary to prevent solidification while the citricacid was added. To convert the free fatty acids into methylesters foranalysis by Gas Chromatography, 0.025 g of the free fatty acids, 5 ml ofa 4% solution of HCl and ethanol were added to a test tube. Nitrogen wasadded to the tube, then the tube was sealed and placed in a water bathat 60° C. for 20 min. The tube was then cooled and 1 ml purified waterand 5 ml isooctane were added. Nitrogen was added to the tube and thetube was shaken for 30 seconds. The resulting upper layer was added to 1μl of purified water in a new test tube and again shaken under nitrogen.The resulting upper layer was then washed of isooctane and decanted intoa third test tube. A small amount of sodium sulfate was added for waterabsorption. A 1 μl sample was then injected directly into the Gaschromatograph.

The gas chromatography conditions were as follows:

System: Perkins-Elmer Auto System Injector: Splitless at 240° C.Detector: Flame Ionization Detector at 280° C. Carrier: Helium Column:WCOT Fused Silica 0.25 mm X100M, CP-SL 88 for FAME, DF 0.2 Oven Program:80° C. (0 min.) increasing to 220° C. at 10° C. per min. and held at220° C. for 10 min.

All results are expressed as the relative peak area percentage.Standards are generally unavailable, so the peaks which eluted wereverified with other systems. GC-MS determines the number, but not theposition of cis and trans bonds. Therefore, NMR analysis was used toverify the bond positions. The main peaks were c9,t11 and t10,c12. ForNMR analysis of CLA isomers, please see Marcel S. F. Lie Ken Jie and J.Mustafa, Lipids, 32 (10) 1019-34 (1997), incorporated herein byreference.

This data, presented in Table 2 and summarized in Table 10, demonstratesthat isomerization of safflower oil using polypropylene glycol as asolvent, KOH as a catalyst, and low temperatures results in theproduction of conjugated linoleic acid lacking 8,10 and 11,13 isomers.The highly polar columns utilized in this experiment may be successfullyused to separate the 8,10 and 11,13 isomers from c9,t11 and t10,c12isomers. The 8,10 isomers tend to coelute or elute just after the c9,t11isomer. The 11,13 isomer elutes in front of the t10,c11 isomer orcoelutes with the t10,c12 isomer, depending on the column conditions.

TABLE 2 Peak Time Component Area Area Height # (Min) Name (%) (μV · s)(μV) 1 38.164 0.08 4101.65 622.28 2 49.539 C16:0 6.29 335897.80 32745.953 53.107 C16:1 0.06 3240.60 447.82 4 61.620 C18:0 2.38 127182.3012999.14 5 64.821 C18:1 c9 12.34 659111.72 52209.40 6 65.254 0.5730402.68 3475.09 7 67.263 0.11 5757.35 758.08 8 67.940 0.10 5523.00700.44 9 68.755 0.24 12816.90 1543.27 10 69.310 0.22 11803.80 1430.59 1169.846 C18:2 c9, c12 0.44 23336.75 2500.24 12 73.618 0.28 14828.701838.66 13 76.621 0.16 8400.65 1050.19 14 77.388 CLA c9, t11 36.511950669.98 124313.83 15 78.370 CLA t10, c12 37.16 1985488.96 132265.3316 78.664 CLA c9, c11 1.06 56583.10 5699.43 17 78.880 CLA c10, c12 1.2667503.55 4572.65 18 80.102 CLA t9, t11/ 0.73 39110.00 4743.28 t10, t1219 85.165 0.03 1621.65 231.32 100.00 5343381.15 384147.01

EXAMPLE 2 Production of CLA with Alcoholate Catalysts

This example describes the production of CLA from safflower oil usingpotassium methylate as a catalyst. Distilled methyl ester of sunfloweroil (41.5 g) was placed in a reactor with 0.207 g methanol and 0.62 gpotassium methylate, and the reactor purged with nitrogen beforeclosing. The contents of the reactor were stirred while to 120° C. Thereaction was then allowed to proceed at 120° C. for 4 hours. the reactorwas then cooled to 80° C. and the contents were transferred to aseparating funnel and washed with hot distilled water and then with hotwater containing citric acid. The methylester was then dried undervacuum with moderate heat. The dried methyl ester was dissolved inisooctane and analyzed by GLC with a Perkin Elmer autosampler. Thecolumn was a highly polar fused silica type. the following program wasused:

Injection: Splitless at 250° C. Detection: Flame ionization detector at280° C. Carrier: Helium at psig. Oven program: 80° C.-130° C. (45°C./min.), then 1° C./min. to 220° C. and 220° C. throughout for 10 min.Column: WCOT FUSED SILICA 0.25 mm 100 m, CP-SIL 88 for FAME, df + 0.2.

The CLA obtained consisted almost exclusively of the c9,t11 and t10,c12isomers of CLA as shown in Table 3.

TABLE 3 CLA Produced by Isomerization with Potassium-Methylate FattyAcid Before Isomerization After Isomerization C 16:0 5.41 5.54 C 18:03.87 3.72 C 18:1 29.01 29.19 C 18:2, c9, c12 59.43 0.84 CLA, c9, c11 028.84 CLA, t10, c12 0 28.45 CLA, c9, c11 0 0.56 CLA, c10, c12 0 0.40CLA, t9, t11; t10, t12 0 0.27

EXAMPLE 3 Production of CLA Powder

This example describes the production of a powder containing CLAacylglycerides of the present invention. The CLA acylglycerides may beprepared as described above. Warm water (538.2 ml at 110-120° F.) andHI-CAP 100 (approximately 230.9 g, National Starch, Bridgewater, N.J.)are combined and agitated until the dispersion is free of any lumps. CLAtriglyceride (230.9 g) is then added and the mixture homogenized for 2min in an Arde Berinco lab homogenizer at setting 30. The pre-emulsionis then homogenized at full speed for 2-5 min (one pass at 3500 psitotal pressure). The particle size is checked and should be from about0.8 to 1.0 microns. The emulsion is then spray dried in a seven footconical dryer at the following settings: inlet temperature (190-215°C.); outlet temperature (95-100° C.). Outlet temperature is maintainedby adjusting the emulsion feed rate. This process produces a freeflowing powder containing approximately 50% CLA triglyceride.

EXAMPLE 4 Preparation of CLA Isomers

This Example describes the production c11,t13 octadecadienoic acid fromt10,c12 octadecadienoic acid. Fifty grams of KOH were dissolved inpropylene glycol under moderate heating. One hundred grams of 98%linoleic acid were then added to the mixture, and the mixture heated to150° C. and stirred for 3 hours. The mixture was then cooled and washedseveral times with hot water and then dried under vacuum at moderateheat. The resulting CLA mixture consisted of c9,t11 and t10,c12octadecadienoic acid as well as traces of CLA isomers. The mixture wasconverted to methylester by reflux boiling in acidic methanol. Fiftygrams of conjugated free fatty acids were dissolved in methanolcontaining 4.5% sulfuric acid and boiled under reflux conditions for 1hour in a water bath. The mixture was cooled and the bottom layerdiscarded. Fresh methanol with 4.5% sulfuric acid was added and themixture boiled for an additional hour under reflux conditions. Aftercooling, this methylester mixture was washed several times with waterand then dried under vacuum at moderate heat. Ten grams of themethylester were dissolved in acetone and cooled overnight to −60° C. ina freezer. A solid precipitate was recovered by filtration andre-dissolved in acetone and again cooled to −60° C. overnight. Theprecipitate was dried under vacuum and shown by GLC analysis to contain97% t10,c12 CLA. The analytical equipment consisted of a Perkin ElmerGLC with auto-sampler. The column was a highly polar fused silica type.The following program setting were used:

Injection: Splitless at 250° C. Detection: Flame Ionization Detector at280° C. Carrier: Helium at psig. Oven program: 80° C.-130° C. (45°C./min), then 1° C./min to 220° C. and 220° C. throughout for 10 min.Column: WCOT FUSED SILICA 0.25 mm × 100 m, CP-SIL 88 for FAME, df = 0.2.

One gram of purified t10,c12 isomer was then covered with nitrogen in asealed tube and heated for two hours at 220° C. After cooling, theresulting methylesters were analyzed by GC as above. The relativecontent of t10,c12 in the mixture was reduced to 52.32% and the c11,t13isomer was present at a level of 41.96% (See Table 4).

TABLE 4 Conversion of t10, c12 isomer to c11, t13 isomer Isomer % Beforeheating % After heating c11, t13 0 41.57 t10, c12 97.34 51.72 C11, c13 01.44 c10, c12 0 2.70 t11, t13 0 0.54 t10, t12 0.7 1.05

EXAMPLE 5 Preparation of CLA Isomers

This Example describes the production t8,c10 octadecadienoic acid fromc9,t11 octadecadienoic acid. Purified c9,t11 octadecadienoic acid may beobtained from commercial sources (Matreya, State College, Pa.) or byfermentation with rumen microorganisms (See, e.g., U.S. Pat. No.5,674,901, incorporated herein by reference). The purified c9,t11octadecadienoic is converted to a high percentage (e.g., 25% to 50%)t8,c10 octadecadienoic acid by placing the c9,t11 octadecadienoic acidin a sealed tube under nitrogen and heating to 220° C. for about 2hours.

EXAMPLE 6 Preparation of CLA Isomers

This Example describes the production t6,c8 octadecadienoic acid fromc7,t9 octadecadienoic acid. Purified c7,t9 octadecadienoic acid may beobtained by preparative scale gas chromatography. The purified c7,t9octadecadienoic is converted to a high percentage (e.g., 25% to 50%)t6,c8 octadecadienoic acid by placing the c9,t11 octadecadienoic acid ina sealed tube under nitrogen and heating to 220° C. for about 2 hours.

EXAMPLE 7 Preparation of CLA Isomers

This Example describes the production c12,t14 octadecadienoic acid fromt11,c13 octadecadienoic acid. Purified t11,c13 octadecadienoic acid maybe obtained by preparative scale gas chromatography (e.g., following theprocess described in Example 1). The purified t11,c13 octadecadienoic isconverted to a high percentage (e.g., 25% to 50%) c12,t14octadecadienoic acid by placing the c9,t11 octadecadienoic acid in asealed tube under nitrogen and heating to 220° C. for about 2 hours.

EXAMPLE 8 Intermediates for Chemical Synthesis of Acylglycerols

The following intermediate compounds are useful for the chemicalsynthesis of triglycerides containing CLA isomers and/or other fattyacids. The use of these intermediates is further described in Examples12-23.

1,3-dipalmitoyl acetone (Intermediate 1)

1,3-dipalmitoyl acetone is prepared from 1,3-dihydroxy acetone (as adimer) and palmitoyl chloride with pyridine as base according to P. H.Bentley et al. in J. Org. Chem. 35:2082 (1970).

1,3-di(c9,t11-octadecadienoyl) acetone (Intermediate 2)

The title compound is prepared from 1,3-dihydroxyacetone andc9,t11-octadecadienoyl acid chloride (prepared from the correspondingacid thionyl chloride or phosphorous penta chloride according tostandard methods).

1,3-di(t10,c12-octadecadienoyl) acetone (Intermediate 3)

The title compound is prepared as intermediate 2 fromt10,c12-octadecadienoid acid and 1,3-dihydroxyacetone.

1,3-palmitoyl glycerol (Intermediate 4)

The title compound is prepared by reduction of 1,3-palmitoyl acetone(Intermediate 1) using sodium borhydride according to P. H. Bentley etal. in J. Org. Chem. 35:2082 (1970).

1,3-di(c9,t11-octadecadienoyl)-glycerol (Intermediate 5)

The title compound is prepared from 1,3-di(c9,t11-octadecadienoyl)acetone (Intermediate 2) according to the method described forIntermediate 4.

1,3-di (t10,c12-octadecadienoyl) glycerol (Intermediate 6)

The title compound is prepared from 1,3-di(t10,c12-octadecadienoyl)acetone (Intermediate 3) according to the method described forIntermediate 4.

1,3-Benzylidene glycerol (Intermediate 7)

1,3-Benzylidene glycerol is prepared from benzaldehyde and anhydrosglycerol as described by Hilbert et al in J. Am. Chem. Soc. 51:1601(1929).

2-Nosyl-1,3-benzylidene glycerol (Intermediate 8)

4-Nitrobenzenesulphonyl chloride (14.7 g, 12 ekv) and benzylideneglycerol (Intermediate 7) are dissolved in dichloromethane (400 ml),Triethylamine (20 ml) and 4-dimethylamino-pyridine (400 mg) are addedand the mixture is stirred at 0° C. for 2 hours and kept at 5° C. for 12hours. Dichloromethane (800 ml) is added and the solution is washed withsaturated aqueous sodium hydrogen carbonate solution (3×200 ml). Theorganic solution is then washed with water (2×200 ml), dried with sodiumsulfate and evaporated to 200 ml. The solution is filtered through aplug of silica gel and evaporated. The title compound is isolated bycrystallization from ethyl acetate.

2-Palmitoyl-1,3-benzylidene glycerol (Intermediate 9)

2-Nosyl-1,3-benzylidene glycerol (Intermediate 8) (2 g, 1 ekv) isdissolved in dimethylformamide (100 ml). The cesium salt of palmitinicacid (1 ekv) (prepared from palmitinic acid and cesium carbonate) isadded, and the reaction mixture is stirred at ambient temperature for 12hours. The mixture is evaporated and the title compound is isolatedafter purification (silica column, ethyl acetate and hexane).

2-(c9,t11-octadecadienoyl)-1,3-benzylidene glycerol (Intermediate 10)

The title compound is prepared as Intermediate 9 usingc9,t11-octadecadienoic acid cesium salt (prepared from the acid andcesium carbonate).

2-(t10,c12-octadecadienoyl)-1,3-benzylidene glycerol (Intermediate 11)

The title compound is prepared as Intermediate 10 usingt10,c12-octadecadienoic acid cesium salt (prepared from thecorresponding acid and cesium carbonate).

2-Palmitoyl-1,3-glycerol (Intermediate 12)

The title compound is prepared by hydrolysis of2-palmitoyl-1,3-benzylidene glycerol (Intermediate 9).2-Palmitoyl-1,3-benzylidene glycerol (2 g, 1 ekv) and boronic acid(0,56, 2 ekv) are suspended in triethylborate (20 ml). The mixture isstirred at ambient temperature until the solution is clear. The mixtureis evaporated. The crude product is dissolved in diethyl ether (100 ml)and washed with water (2×50 ml). The organic phase is dried with sodiumsulfate and evaporated yielding the title compound.

2-(c9,t11-octadecadienoyl)-1,3 glycerol (Intermediate 13)

The title compound is prepared by hydrolysis of2-c9,t11-octadecadienoyl-1,3-benzylidene glycerol (Intermediate 10)according to the method in Intermediate 12.

2-(t10,c12-octadecadienoyl)-1,3-glycerol (Intermediate 14)

The title compound is prepared by hydrolysis of2-t10,c12-octadecadienoyl-1,3-benzylidene glycerol (Intermediate 11)according to the method in Intermediate 12.

1,3-Ditosyl-2-palmitoyl-1,3-glycerol (Intermediate 15)

2-Palmitoyl-1,3-glycerol (Intermediate 12) (0.4 g, 1 ekv) is dissolvedin pyridine (10 ml) p-Toluenesulphonyl chloride (0.5 g, 2.5 ekv) isgradually added. The mixture is stirred for 12 hours at ambienttemperature, evaporated and the title compound is crystallized fromethanol.

1,3 -Ditosyl-2-(c9,t11-octadecadienoyl) glycerol (Intermediate 16)

The title compound is prepared similar to Intermediate 15 starting withIntermediate 12.

1,3-Ditosyl-2-(t10,c12-octadecadienoyl) glycerol (Intermediate 17)

The title compound is prepared similar to Intermediate 15 starting withIntermediate 14.

1,3-Dipalmitoyl-2-nosyl-glycerol (Intermediate 18)

1,3-palmitoyl glycerol (Intermediate 4) (2.5 g, 4.4 mmol) is dissolvedin dichloromethane (60 ml), 4-nitrobenzenesulphonyl chloride (1.2 g, 5.5mmol) 4-dimethylaminopyridine (25 mg, 0.2 mmol) and triethyl amine (1.12g, 11 mmol) are added. The mixture is stirred for 3 days at 0° C.,Dichloromethane (200 ml) is added and the organic solution is washedwith saturated aqueous sodium hydrogen carbonate solution (2×200 ml) andthen washed with water (2×200 ml). The organic phase is dried withsodium sulfate and evaporated. The title compound is purified bychromatography (silica, ethyl acetate and hexane).

1,3-di (c9,t11-octadecadienoyl)-2-nosyl glycerol (Intermediate 19)

The title compound is prepared similar to Intermediate 18 starting fromIntermediate 5.

1,3-di(t10,c12-octadecadienoyl)-2-nosyl glycerol (Intermediate 20)

The title compound is prepared similar to Intermediate 18 starting fromIntermediate 6.

EXAMPLE 9 1,3-di(c9,t11-octadecadienoyl)-2-palmitoyl glycerol(triglyceride)

1,3-Ditosyl-2-palmitoyl-1,3-glycerol (Intermediate 15) (1 ekv) andc9,t11-octadecadienoic acid cesium salt (2 ekv) are suspended in tolueneand dimethylformamide (1:1). The mixture is stirred at ambienttemperature for 12 hours. The solvents are removed by evaporation andthe title compound is isolated after chromatography (silica, ethylacetate and hexane).

EXAMPLE 10 1,3-di(t10,c12-octadecadienoyl)-2-palmitoyl glycerol(triglyceride)

The title compound is prepared similar to Example 9 starting witht10,c12-octadecadienoic acid cesium salt.

EXAMPLE 11 1,2,3-tri(c9,t11-octadecadienoyl)-glycerol (triglyceride)

The title compound is prepared similar to Example 9 starting withIntermediate 16 and c9,t11-octadecadienoic acid cesium salt. Thiscompound can also be prepared directly from glycerol and activated acidderivative (acid chloride or acid anhydride) or by use of couplingagents like for example dicyclohexyl-carbodiimide (DCC).

EXAMPLE 121,3-di(t10,c12-octadecadienoyl-2-(c9,t11-octadecadienoyl)-glycerol(triglyceride)

The title compound is prepared similar to Example 9 starting withIntermediate 16 and t10,c12-octadecadienoic acid cesium salt.

EXAMPLE 13 1,3-Dipalmitoyl-2-(c9,t11-octadecadienoyl)-glycerol(triglyceride)

The title compound is prepared similar to Example 9 starting withIntermediate 16 and palmitoyl cesium salt.

EXAMPLE 14 1,2,3-tri(t10,c12-octadecadienoyl)-glycerol (triglyceride)

The title compound is prepared similar to Example 9 starting withIntermediate 17 and t10,c12-octadecadienoic acid cesium salt. Thiscompound can also be prepared directly from glycerol and activated acid.

EXAMPLE 151,3-di(c9,t11-octadecadienoyl)-2-(t10,c12-octadecadienoyl)-glycerol(triglyceride)

The title compound is prepared similar to Example 9 from Intermediate 17and c9,t11-octadecadienoid acid cesium salt.

EXAMPLE 16 1,3-dipalmitoyl-2-(t10,c12-octadecadienoyl)-glycerol(triglyceride)

The title compound is prepared similar to Example 9 from Intermediate 17and palmitoyl cesium salt.

EXAMPLE 17 1,3-Diplamitoyl-2-(c9,t11-octadecadienoyl)-glycerol(triglyceride)

Same compound as in Example 13, but different method.1,3-Dipalmitoyl-2-nosyl glycerol (Intermediate 18) (0.25 g, 0.3 mmol) isdissolved in dimethylformamide (5 ml). c9,t11-Octadecadineoic acidcesium salt (1 ekv) is added and the mixture is stirred for 48 hours atambient temperature. Dichloromethane (50 ml) is added and the mixture iswashed with saturated aqueous sodium hydrogen carbonate solution (2×30ml). The organic phase is dried (Na₂SO₄) and evaporated—The titlecompound is isolated after chromatography (silica, ethyl acetate andhexane).

EXAMPLE 18 1,3-Dipalmitoyl-2-(t10,c12-octadecadienoyl)-glycerol(triglyceride)

This compound is the same as in Example 16, but different method. Thetitle compound is prepared similar to Example 20 starting fromIntermediate 18 and t10,c12-octadecadienoic acid cesium salt.

EXAMPLE 19 1,2,3-tri(c9,t11-octadecadienoyl)-glycerol (triglyceride)

Same compound as Example 11, but different method. This compound isprepared similar to Example 20 starting from Intermediate 19 andc9,t11-octadecanoic acid cesium salt.

EXAMPLE 20 1,3-di(t10,c12-octadecadienoyl)-2-palmitoyl glycerol(triglyceride)

Same compound as Example 10, but different method. This compound isprepared similar to Example 17 starting from Intermediate 20 andpalmitic acid cesium salt.

In these examples, palmitic acid has been utilized as a representativenon-CLA fatty acid. Similar triglycerides can be prepared from otherfatty acids including both saturated, unsaturated and poly-unsaturatedfatty acids. CLA triglycerides conjugates can also be prepared withbiologically active acids or drugs. Additionally, mixtures of CLAs canbe used to prepare mixtures of triglycerides.

What should be clear from above is that the present invention providesnovel acylglycerides comprising CLA and at least one other fatty acylresidue (e.g., ω3, ω6, and ω9 fatty acyl residues) which can be used inpharmaceutical compositions, animal feeds and in products suitable forhuman consumption. All publications and patents mentioned in the abovespecification are herein incorporated by reference. Variousmodifications and variations of the described method and system of theinvention will be apparent to those skilled in the art without departingfrom the scope and spirit of the invention. Although the invention hasbeen described in connection with specific preferred embodiments, itshould be understood that the invention as claimed should not be undulylimited to such specific embodiments. Indeed, various modifications ofthe described modes for carrying out the invention which are obvious tothose skilled in medicine, biochemistry, or related fields are intendedto be within the scope of the following claims.

1. An acylglyceride having the following structure:

wherein R1 and R3 are acyl residues selected from the group consistingof 10,12; 9,11; 8,10; and 11,13 octadecadienoate and R2 is selected fromthe group consisting of medium chain fatty acyl residues.
 2. Theacylglyceride of claim 1, wherein said medium chain fatty acyl residueis selected from the group consisting of acyl residues of the followingacids: decanoic acid, undecanoic acid, 10-undecanoic acid, lauric acid,cis-5-dodecanoic acid tridecanoic acid, myristic acid, and myristoleicacid.
 3. A powder comprising at least one type of acylglyceride as setforth in claim
 1. 4. An oil comprising at least one type ofacylglyceride as set forth in claim
 1. 5. The oil of claim 4, whereinsaid oil further comprises an antioxidant.
 6. A food compositioncomprising at least one type of acylglyceride as set forth in claim 1.7. The food composition of claim 6, wherein said food composition is afunctional food, nutritional supplement food, infant food, pregnancyfood, or elderly food.
 8. A pharmaceutical composition comprising atleast one type of glyceride as set forth in claim
 1. 9. A nutritional orpharmaceutical composition comprising at least one type of acylglycerideas set forth in claim 1 and a carrier suitable for oral,intraintestinal, or parenteral administration.